Research Projects

Plants are constantly exposed to herbivory, which results in significant agricultural losses, accounting for over 20% of global crop yield reductions annually. In response, plants have evolved complex defense mechanisms, with jasmonic acid (JA) serving as a key hormone mediating both local and systemic responses to wounding. However, the mechanisms underlying the long-distance transmission of defense signals and the integration of these signals into systemic JA biosynthesis remain poorly understood. Recent work in Arabidopsis thaliana has identified the plasma membrane H⁺-ATPase AHA1 as a critical regulator of wound-induced electrical signaling and systemic JA responses (Kumari et al., 2019). This project is driven by the hypothesis that AHA1 functions as a signaling hub regulated through C-terminal phosphorylation, and that its activity in specific vascular tissues governs the propagation of electrical signals and hormonal responses required for systemic defense activation. Furthermore, precise modulation of AHA1 through post-translational modifications could uncouple defense activation from growth penalties, offering opportunities for translational crop improvement. The project is organized around four scientific objectives: Investigate the role of C-terminal phosphorylation and tissue-specific activity of AHA1 in systemic defense. Using a constitutively active AHA1 variant (AHA1p-AHA1∆C-Venus) expressed under native and vascular-specific promoters, we will evaluate its influence on electrical signaling, JA accumulation, and herbivore resistance. Subcellular localization will be assessed using a VENUS-tagged version through confocal microscopy. Identify key phosphorylation sites in AHA1 involved in defense signaling. Conserved motifs in the C-terminal domain will be aligned and analyzed in silico to predict regulatory phosphosites. Phosphomimic (active/inactive) mutants will be generated and structurally evaluated via ∆∆G calculations. Phosphoproteomic analysis under control and herbivory conditions will validate in vivo phosphorylation events. Functionally characterize AHA1 phospho-mutants in plant defense. Site-directed mutagenesis will generate AHA1 variants for yeast complementation, Arabidopsis transformation, and herbivore bioassays. JA pathway gene expression, electrical signaling profiles, and resistance to Spodoptera spp. will be examined. Candidate kinases and phosphatases will be identified and tested for regulatory roles. Assess the impact of AHA1 regulation on reproductive development and yield. Complemented and mutant lines will be analyzed for floral structure, pollen viability, silique formation, and seed set under both normal and stress conditions. AHA1 expression in reproductive tissues will be assessed using promoter-GUS fusions and qRT-PCR. Achieving the project objectives will provide a mechanistic understanding of how proton pumps regulate systemic defense responses via electrical signaling and JA biosynthesis. By identifying key phosphorylation sites and determining cell-specific requirements for AHA1 function, this research will uncover new regulatory layers in plant defense. Importantly, it opens the possibility for rational design of small molecules or genetic modifications that selectively activate defense without growth trade-offs. This work will significantly advance the field of plant stress biology and contribute to the development of pest-resistant, high-yielding crop varieties in a sustainable manner.